本發(fā)明屬于生物技術(shù)領(lǐng)域，具體涉及一種大腸桿菌-枯草芽孢桿菌高效穿梭表達(dá)載體及其構(gòu)建方法與應(yīng)用。

背景技術(shù)：

枯草芽孢桿菌（Bacillus subtilis）是一種極具潛力的外源蛋白表達(dá)宿主菌，不論是用來高效表達(dá)外源蛋白還是當(dāng)做口服疫苗和全細(xì)胞酶催化劑的載體，都有著許多其他細(xì)菌無可比擬的優(yōu)勢，是研究較多、應(yīng)用廣泛的生物安全菌。B. subtilis特有的生物安全性、遺傳操作簡單、可廉價大規(guī)模培養(yǎng)及芽孢具有的較強(qiáng)的抗逆性，使得B. subtilis的基因工程菌產(chǎn)品，特別是一些工業(yè)酶制劑如α-淀粉酶、蛋白酶、堿性磷酸酶等，已占據(jù)了世界工業(yè)酶制劑產(chǎn)量的一半以上；還有一部分抗生素、維生素、肌苷、核黃素等也已進(jìn)入商業(yè)化生產(chǎn)。從轉(zhuǎn)錄模式上分類，B. subtilis的啟動子可分為組成型啟動子和誘導(dǎo)型啟動子。常見的組成型啟動子有P43啟動子，誘導(dǎo)型啟動子有sacB啟動子、麥芽糖啟動子Pglv、Pspac啟動子、PxylR啟動子。PxylR啟動子中編碼木糖異構(gòu)酶（XylA）的基因序列和木酮糖激酶（XylB）的基因序列構(gòu)成了B. subtilis木糖代謝途徑的重要調(diào)操縱子-木糖操縱子（xyl操縱子）。一般木糖操縱子片段包括整個xylR編碼區(qū)序列，PxylA、PxylB、xylO、XylA前的CRE（catabolite-responsive element）區(qū)和SD序列區(qū)。將外源基因的編碼序列替代XylA，利用木糖代謝途徑中木糖與xylR的相互作用，誘導(dǎo)控制PxylA啟動子表達(dá)外源蛋白。

外源基因轉(zhuǎn)入B. subtilis菌體內(nèi)需要有載體的參與，主要是質(zhì)粒載體。B. subtilis使用的第一代質(zhì)粒載體主要來自其他革蘭氏陽性細(xì)菌，如：pUB110、pHTZ21和pLS11等，第二代使用于B. subtilis的質(zhì)粒載體是穿梭質(zhì)粒，由第一代非B. subtilis的革蘭氏陽性菌天然質(zhì)粒與E. coli質(zhì)粒（一般為pBR322或其衍生的質(zhì)粒）構(gòu)成的載體，第三代質(zhì)粒載體是由B. subtilis的隱性質(zhì)粒與E. coli質(zhì)粒構(gòu)成的嵌合載體，如：pHP3、pMTLBS72、pHT01和pHT43。pHP3由質(zhì)粒pTA1060的復(fù)制子片段、質(zhì)粒pE194的紅霉素抗性基因（Em^r）、質(zhì)粒pC194的頭孢菌素抗性基因（Cm^r）和E. coli質(zhì)粒pUC9的復(fù)制子片段組成，在E. coli和B. subtilis中都能表現(xiàn)所有的抗生素抗性，而且具有較高的B. subtilis轉(zhuǎn)化效率和穩(wěn)定的遺傳性。質(zhì)粒pMTLBS72由來自野生B. subtilis的質(zhì)粒的復(fù)制子片段和E. coli質(zhì)粒pMTL21C組合構(gòu)建，具有較好遺傳穩(wěn)定性，以此為基礎(chǔ)添加強(qiáng)啟動子構(gòu)建成功的質(zhì)粒pHT01，pHT43是已經(jīng)用于生產(chǎn)的商品質(zhì)粒，在連續(xù)傳代和發(fā)酵罐培養(yǎng)中都比較穩(wěn)定。

隨著基因工程的飛速發(fā)展，作為引導(dǎo)蛋白質(zhì)分泌的信號肽也倍受關(guān)注。外源蛋白質(zhì)在宿主菌，如大腸桿菌中的表達(dá)形式多為細(xì)胞內(nèi)不溶性表達(dá)（包涵體），少數(shù)為細(xì)胞外分泌表達(dá)。利用信號肽的特性來引導(dǎo)外源蛋白質(zhì)定位分泌到細(xì)胞特定區(qū)間，提高可溶性，可避免因包涵體復(fù)性帶來的困難。目前研究采用的信號肽基本來自表達(dá)系統(tǒng)自身的信號序列或外源信號序列，或兩者兼而有之。研究表明，多種外源基因連上信號肽后，在原核表達(dá)系統(tǒng)，如大腸桿菌、L型細(xì)菌、乳酸桿菌和芽孢桿菌等中都得到了分泌表達(dá)。信號肽因其特有的功能在外源基因的分泌表達(dá)上被廣泛應(yīng)用。

B. subtilis表達(dá)系統(tǒng)是繼E. coli表達(dá)系統(tǒng)之后研究最為成熟的原核表達(dá)系統(tǒng)。B. subtilis作為外源蛋白分泌表達(dá)的宿主菌具有很大發(fā)展?jié)摿?。利用誘導(dǎo)方便，表達(dá)效率高的啟動子構(gòu)建穩(wěn)定遺傳的穿梭表達(dá)載體是實現(xiàn)外源蛋白高效表達(dá)的有效途徑。因此，構(gòu)建一個便于誘導(dǎo)、表達(dá)效率高和遺傳操作簡便的E. coli-B. subtilis穿梭表達(dá)載體，為今后研究更高效的穿梭表達(dá)載體奠定基礎(chǔ)，對研制枯草芽孢桿菌工程菌的實驗研究和工業(yè)生產(chǎn)的日益迫切的需求提供了更為廣闊的前景。

技術(shù)實現(xiàn)要素：

在本發(fā)明的目的是構(gòu)建一個高效的含有木糖啟動子的E. coli-B. subtilis穿梭表達(dá)載體和一個帶有信號肽的分泌型穿梭表達(dá)載體并對其效率進(jìn)行評估，以滿足實驗研究和工業(yè)生產(chǎn)的日益迫切的需求。

一種高效穿梭表達(dá)載體，所述表達(dá)載體為pHTL，其核苷酸序列如序列表SEQ ID NO:1所示。

所述高效穿梭表達(dá)載體的構(gòu)建方法為：

（1）獲得木糖啟動子序列PxylR和大腸桿菌表達(dá)載體pET-21b中T7表達(dá)區(qū)序列的融合片段PxylR-T7；

所述表達(dá)區(qū)序列包括T7短肽，多克隆位點區(qū)，T7終止子；

（2）融合片段PxylR-T7插入經(jīng)酶切修飾后的骨架載體pHT315的多克隆位點之間。

一種重組表達(dá)載體，其在上述高效穿梭表達(dá)載體的多克隆位點之間插入目的基因。

所述重組表達(dá)載體為pHTL-gfp，pHTL-gus和pHTL-lipa，其序列如SEQ ID NO:14、SEQ ID NO:15和SEQ ID NO:16所示。

包含上述重組表達(dá)載體的工程菌。

所述工程菌為B8-pHTLgfp，B8-pHTLgus和B8-pHTLlipa，含有上述重組表達(dá)載體pHTL-gfp，pHTL-gus，pHTL-lipa。

所述表達(dá)載體在基因gfp，gus和lipa表達(dá)中的應(yīng)用，表達(dá)蛋白為GFP，GUS和LipA。

進(jìn)一步的，所述步驟（1）的方法為：以B. subtilis 168菌株基因組DNA為模板，用引物對xyl-5/xyl-3擴(kuò)增木糖啟動子PxylR序列，大小為1473bp；以大腸桿菌表達(dá)載體pET-21b質(zhì)粒為模板，用引物對T7-5/T7-3擴(kuò)增得到無啟動子的pET-21b中T7表達(dá)區(qū)片段，大小為213bp，其中包括T7短肽、多克隆位點區(qū)（MCS）、T7終止子；回收兩個PCR產(chǎn)物，不添加引物，利用PrimeSTAR Max DNA Polymerase進(jìn)行互補(bǔ)延伸，形成融合片段的中間產(chǎn)物，以此為模板，用引物對xyl-5/T7-3進(jìn)行融合PCR的擴(kuò)增，獲得木糖啟動子PxylR片段和載體pET-21b的T7表達(dá)區(qū)的融合片段PxylR-T7，大小為1686bp，所述骨架載體的pHT315經(jīng)EcoR /Hind 雙酶切后，用Klenow酶和Fastap酶先后進(jìn)行黏末端補(bǔ)平反應(yīng)以及對補(bǔ)平的片段進(jìn)行去磷酸化處理；

所述xyl-5序列如序列表SEQ ID NO:2所示，所述xyl-3序列如SEQ ID NO:3所示，

所述T7-5序列如SEQ ID NO:4所示，所述T7-3序列如SEQ ID NO:5所示。

一種分泌型穿梭表達(dá)載體，將來自納豆芽孢桿菌堿性蛋白酶信號肽序列取代T7表達(dá)區(qū)序列插入上述載體pHTL的木糖啟動子PxylR和多克隆位點之間。

所述分泌型穿梭表達(dá)載體的序列如序列表SEQ ID NO:17所示。

所述分泌型穿梭表達(dá)載體的構(gòu)建方法為：

1）獲得木糖啟動子PxylR部分序列和納豆芽孢桿菌堿性蛋白酶信號肽序列Apre的融合片段PxylRstuI-Apre；

2）插入經(jīng)酶切后的骨架載體pHTL的PxylR中Stu I位點和多克隆位點的BamH I之間。

進(jìn)一步的，所述步驟1）的方法為：以質(zhì)粒pHTL為模板，用引物對xyls-5/xyls-3擴(kuò)增木糖啟動子PxylR部分序列PxylRstuI，大小為1167bp；以納豆芽孢桿菌基因組為模板，用引物對apre-5/apre-3擴(kuò)增得到納豆芽孢桿菌中編碼信號肽的DNA區(qū)段Apre，大小為96bp；回收兩個PCR產(chǎn)物，不添加引物，利用PrimeSTAR Max DNA Polymerase進(jìn)行互補(bǔ)延伸，形成融合片段的中間產(chǎn)物，以此為模板，用引物對xyls-5/apre-3進(jìn)行融合PCR的擴(kuò)增，獲得木糖啟動子PxylRstuI和納豆芽孢桿菌信號肽表達(dá)區(qū)的融合片段PxylRstuI-Apre，大小為1263bp，所述骨架載體的pHTL經(jīng)Stu I/BamH I雙酶切；

所述xyls-5序列如SEQ ID NO:18所示，所述xyls-3序列如SEQ ID NO:19所示，

所述apre-5序列如SEQ ID NO:20所示，所述apre-3序列如SEQ ID NO:21所示。

信號肽插入PxylR和多克隆位點之間的方法和限制性內(nèi)切酶的選擇：在PxylR和Apre相接處通過引物設(shè)計引入限制性內(nèi)切酶PmlI的酶切位點CAC|GTG，在Apre和多克隆位點之間通過PCR引物設(shè)計引入了限制性內(nèi)切酶Ngo MIV酶切位點序列G|CCGGC；同時在信號肽末端和Ngo MIV之間引入了有利于信號肽酶酶切的氨基酸序列AlaGluAla，其編碼序列為：“GCGCAGGCT”。

所述分泌表達(dá)載體上替換Apre信號肽序列的策略：通過擴(kuò)增信號肽的引物引入上述酶切位點，即5’端引物引入Pml I酶切位點、3’端引物引入Ngo MIV酶切，通過上述一對引物擴(kuò)增得到的任何信號肽的DNA編碼區(qū)可以通過這兩個酶切位點插入pHTL的PxylR和多克隆位點之間，以替換Apre信號肽并提高多克隆位點處插入外源蛋白表達(dá)效率和信號肽切除效率。

一種重組表達(dá)載體，在上述分泌型穿梭表達(dá)載體的多克隆位點之間插入目的基因。

所述重組表達(dá)載體分別為pHTZ-gus，pHTZ-tgl1，pHTZ-lipa，其序列如SEQ ID NO:22、SEQ ID NO:23、SEQ ID NO:24所示。

包含上述重組表達(dá)載體的工程菌。所述工程菌為B8-pHTZgus，B8-pHTZtgl1和B8-pHTZlipa，含有重組表達(dá)載體pHTZ-gus，pHTZ-tgl1，pHTZ-lipa。

所述表達(dá)載體在基因gus，tgl1和lipa蛋白中的應(yīng)用，表達(dá)蛋白為GUS，Tgl1和LipA。

本發(fā)明的有益效果：本發(fā)明是在E. coli-B. subtilis穿梭載體pHT315的基礎(chǔ)上構(gòu)建，通過將枯草芽孢桿菌Bs168的木糖啟動子序列與來自E. coli表達(dá)載體pET21b的T7表達(dá)區(qū)序列融合，構(gòu)建了一個高效穿梭表達(dá)載體pHTL，以gfp，gus和lipa為報告基因，檢測載體的表達(dá)能力和遺傳穩(wěn)定性；同時，將來自納豆芽孢桿菌的堿性蛋白酶信號肽序列代替載體pHTL上的T7肽編碼區(qū)，構(gòu)建成高效分泌型穿梭表達(dá)載體pHTZ，以gus，tgl1和lipa為報告基因，檢測載體的表達(dá)能力，兩種載體均可以正確表達(dá)相應(yīng)蛋白并具有生物學(xué)活性，具有廣泛的應(yīng)用前景。

附圖說明

圖1 為pHTL載體構(gòu)建示意圖。

圖2 為pHTL載體誘導(dǎo)表達(dá)區(qū)構(gòu)建示意圖。

圖3 為pHTL載體PCR和酶切鑒定；

圖中，1：1 kb DNA Ladder Marker；2：質(zhì)粒pHTL的EcoR I單酶切；3：質(zhì)粒pHTL 的Not I單酶切；4：質(zhì)粒pHTL的Xho I單酶切；5：質(zhì)粒pHTL的SmaI單酶切；6：質(zhì)粒pHTL的Kpn I單酶切；7：質(zhì)粒pHTL的PCR產(chǎn)物。

圖4：重組質(zhì)粒pHTL-gfp，pHTL-gus和pHTL-lipa酶切鑒定；

圖中，1，5，6：1 kb DNA Ladder Marker； 2：質(zhì)粒pHTL-gfp雙酶切；3：100 bp DNA Ladder Marker；4：質(zhì)粒：pHTL-gus雙酶切；7：質(zhì)粒:pHTL-lipa雙酶切。

圖5 為菌株B8-pHTLgfp的熒光顯微鏡觀察結(jié)果；

A：WB800可見光下觀察；B：WB800紫外光下觀察；C：B8-pHTLgfp可見光下觀察；D：B8-pHTLgfp紫外光下觀察。

圖6 為菌株B8-pHTLgus的顏色變化觀察結(jié)果；

圖中，A：WB800；B：B8-pHTLgus。

圖7 為菌株B8-pHTLgfp的質(zhì)粒穩(wěn)定性分析。

圖8 為pHTZ載體構(gòu)建示意圖。

圖9 為pHTZ載體表達(dá)區(qū)構(gòu)建示意圖。

圖10 為pHTZ載體PCR和酶切鑒定；

圖中，1，3，8：1 kb DNA Ladder Marker；2：質(zhì)粒pHTZ 的Stu I/BamH I雙酶切；4：質(zhì)粒pHTZ的PCR產(chǎn)物；5：質(zhì)粒pHTZ的Xho I單酶切；6：質(zhì)粒pHTZ的Sal I單酶切；7：質(zhì)粒pHTZ的Hind III單酶切。

圖11 為重組質(zhì)粒pHTZ-gus，pHTZ-tgl1，pHTZ-lipa酶切鑒定；

圖中，1，5：1 kb DNA Ladder Marker；4：100 bp DNA Ladder Marker；2：質(zhì)粒pHTZ-gus雙酶切產(chǎn)物；3：質(zhì)粒pHTZ-lipa雙酶切產(chǎn)物；6：質(zhì)粒pHTZ-tgl1雙酶切產(chǎn)物。

圖12 為菌株B8-pHTZgus的顏色變化觀察結(jié)果；

圖中，A：B8-pHTZgus；B：WB800。

圖13 為菌株B8-pHTZtgl1產(chǎn)酶蛋白的量與BSA交聯(lián)程度的SDS-PAGE分析；

圖中，A：WB800；B：B8-pHTZtgl1。

具體實施方式

下面結(jié)合附圖和具體實施例對本發(fā)明做進(jìn)一步說明。

實施例1

（1）B. subtilis168基因組提取

無菌條件下，挑取B. subtilis 168單菌落，接種于LB培養(yǎng)基中，37 ℃，200 rpm培養(yǎng)12 h；按2%接種量接種到20 mL的LB培養(yǎng)基中37 ℃，200 rpm過夜培養(yǎng)；將菌液收集在1.5 mL離心管中，12000 rpm離心1 min，收集所有菌體，用TE洗一次；加入200 μL TE緩沖液震蕩混勻；加入200 μL 2%的SDS緩沖液混勻；加入400 μL異硫氰酸胍已酸鈉混合液；12000 rpm離心5 min，將上清轉(zhuǎn)移到一個新的離心管中，加入600 μL異丙醇混勻；12000 rpm離心10 min去上清，加入500 μL 70%乙醇混勻；12000 rpm離心10 min，倒去上清，室溫晾干至無乙醇?xì)馕叮患尤?0 μL滅菌的去離子水，加入5 μLRNA酶溶液，37 ℃孵育30 min，用1%的瓊脂糖凝膠進(jìn)行電泳，檢測提取的的基因組DNA。

（2）融合片段擴(kuò)增

分別以B. subtilis 168基因組DNA為模板用引物對xylr-5/xylr-3（SEQ ID NO:2/ SEQ ID NO:3）擴(kuò)增木糖1473bp的啟動子PxylR序列；以大腸桿菌表達(dá)載體pET-21b質(zhì)粒為模板，用引物對T7-5/T7-3（SEQ ID NO:4/ SEQ ID NO:5）擴(kuò)增得到213bp的無啟動子的pET-21b中T7表達(dá)區(qū)片段，其中包括多克隆位點區(qū)；回收兩個PCR產(chǎn)物，不添加引物，利用PrimeSTAR Max DNA Polymerase進(jìn)行互補(bǔ)延伸，形成融合片段的中間產(chǎn)物，以此為模板，用引物對xylr-5/T7-3進(jìn)行融合PCR的擴(kuò)增，獲得木糖啟動子序列PxylR和載體pET-21b表達(dá)區(qū)的融合片段PxylR-T7，大小為1686bp。

（3） 質(zhì)粒pHT315平端化

質(zhì)粒pHT315使用限制性內(nèi)切酶EcoR 和Hind 雙酶切，將回收的片段用klenow酶進(jìn)行黏末端補(bǔ)平反應(yīng)，利用Fastap酶對補(bǔ)平的片段進(jìn)行去磷酸化處理，電泳回收去磷酸化的質(zhì)粒pHT315酶切補(bǔ)平片段。

（4）穿梭表達(dá)載體pHTL的構(gòu)建

將融合片段PxylR-T7用限制性內(nèi)切酶Sam I進(jìn)行單酶切，回收后的片段與質(zhì)粒pHT315酶切補(bǔ)平去磷酸化得到的片段連接，得到重組質(zhì)粒pHTL。該質(zhì)粒含有PxylR序列和T7肽（不含啟動子）、多克隆位點區(qū)、T7終止子，物理圖譜如圖2。重組質(zhì)粒經(jīng)多種限制性內(nèi)切酶單酶切、PCR及測序鑒定（SEQ ID NO:6/ SEQ ID NO:7），證明構(gòu)建正確圖3，序列見（SEQ ID NO:1）。

（5） 新型高效穿梭表達(dá)載體的效率評估

以質(zhì)粒pMD-19T-gfp為模板，引物對5F-H/3R-H（SEQ ID NO:8/ SEQ ID NO:9）進(jìn)行PCR反應(yīng)擴(kuò)增大小為756bp的gfp基因；以質(zhì)粒pBI121為模板，引物對gusp-5/gusp-3（SEQ ID NO:10/ SEQ ID NO:11）進(jìn)行PCR反應(yīng)擴(kuò)增大小為1812bp的gus基因；以質(zhì)粒pMD-19lipa為模板，引物對LSal-5/Lhind-3（SEQ ID NO:12/ SEQ ID NO:13）進(jìn)行PCR反應(yīng)擴(kuò)增大小為1842 bp的lipa基因。分別用限制性內(nèi)切酶EcoR 和Hind 、Hind和Xho、Sal和Sph I雙酶切回收的gfp、gus和lipa基因，導(dǎo)入pHTL的相應(yīng)酶切位點間，轉(zhuǎn)化E. coli JM109，得到重組質(zhì)粒pHTL-gfp，pHTL-gus和pHTL-lipa，經(jīng)酶切測序鑒定正確（圖4），證明構(gòu)建正確，序列見（SEQ ID NO:14、SEQ ID NO:15、SEQ ID NO:16）。再轉(zhuǎn)入E. coli SCS110去甲基化后，采用熱擊轉(zhuǎn)化的方法將重組質(zhì)粒導(dǎo)入B.subtilis WB800感受態(tài)細(xì)胞中，獲得工程菌B8-pHTLgfp，B8-pHTLgus和B8-pHTLlipa。

工程菌B8-pHTLgfp中GFP的表達(dá)：挑選重組工程菌單菌落接種于LB液體培養(yǎng)基中培養(yǎng)過夜，離心收集細(xì)菌，棄去上清，用滅菌的生理鹽水洗滌2次。挑去菌體涂在載玻片上在熒光顯微鏡下觀察。用滅菌生理鹽水稀釋菌體至OD為1，用熒光分光光度計掃描適合檢測gfp基因表達(dá)的發(fā)射波長和激發(fā)波長。結(jié)果如圖5，熒光分光光度計掃描菌體的最佳激發(fā)波長為390 nm，發(fā)射波長為507 nm，說明pHTL載體可以正確表達(dá)gfp基因。

工程菌B8-pHTLgus中GUS表達(dá)檢測：挑取工程菌單菌落接種于5 mL含有紅霉素(10 μg /mL) LB液體培養(yǎng)基中，活化后轉(zhuǎn)接新的液體培養(yǎng)基中震蕩培養(yǎng)至菌體OD為0.1時，加入終濃度為1%的D-Xylose，誘導(dǎo)8 h后收獲。取1 mL培養(yǎng)基，離心去掉上清收集菌體，滅菌的生理鹽水洗滌2次，保留菌體。用450 μLGUS基液重懸菌體，加入50 μL的X-Gluc母液，混勻。37 ℃保溫，5 min后肉眼觀察染色結(jié)果（圖6）。證明載體pHTL能成功的高效表達(dá)GUS蛋白。

工程菌B8-pHTLlipa中脂肪酶的表達(dá)：建立對硝基苯酚（p-NP）的濃度標(biāo)準(zhǔn)曲線。將活化的工程菌B8-pHTLlipa轉(zhuǎn)接LB液體培養(yǎng)基中震蕩培養(yǎng)，當(dāng)菌體OD為0.1時，加入終濃度為1%的D-Xylose，誘導(dǎo)8 h后收獲。離心收集菌體，用無菌生理鹽水洗2遍。甩干水分，測菌體干重。同時，收獲以同樣接種培養(yǎng)同時期收獲但不加D-Xylose的工程菌WB800-lipa和以同樣接種培養(yǎng)同時期收獲的B. subtilis WB800菌體，作為脂肪酶表達(dá)的參照。用細(xì)胞破碎液重懸待測的菌體，加入溶菌酶溶液（100 mg/mL），37 ℃保溫30 min，用細(xì)胞破碎儀進(jìn)行菌體破碎，收集上清液進(jìn)行酶活測定。根據(jù)建立p-NP濃度與410 nm下吸光值的標(biāo)準(zhǔn)曲線，計算破碎液中的p-NP量，根據(jù)脂肪酶活力定義計算收獲菌體的脂肪酶的酶活（U/mg），作為對照的WB800幾乎無脂肪酶活性（0.25±0.07 U/mg），同時，無木糖誘導(dǎo)的工程菌WB800-lipa的菌體酶活力為0.34±0.05 U/mg.，誘導(dǎo)后的脂肪酶活力為72.73±0.25 U/mg，提高了近214倍，說明載體pHTL成功并高效表達(dá)了LipA蛋白。

質(zhì)粒穩(wěn)定性檢測：接種重組工程菌B8-pHTLgfp于含有紅霉素的LB液體培養(yǎng)基中培養(yǎng)活化轉(zhuǎn)接至LB液體培養(yǎng)基中搖瓶培養(yǎng) 12 h 后，以1%的接種量轉(zhuǎn)入新的LB液體性培養(yǎng)基中（不含紅霉素），開始連續(xù)搖瓶培養(yǎng)試驗。每12 h按1%接種量轉(zhuǎn)接一次為一代工轉(zhuǎn)接10代。 每一代取樣1 mL進(jìn)行平板稀釋涂布檢測質(zhì)粒的缺失程度。檢測時，取菌液用無菌水逐級稀釋，取三個合適的梯度，分別涂在含有和不含有紅霉素的固體LB培養(yǎng)基平板上，每個處理三個重復(fù)。 倒置在 37℃培養(yǎng)箱中培養(yǎng)24h，計算各皿中的菌落數(shù)。計算出質(zhì)粒的穩(wěn)定率（穩(wěn)定率=含有抗生素平板的菌落數(shù)/不含有抗生素平板的菌落數(shù)）。結(jié)果（圖8）表明在無抗生素選擇壓力下質(zhì)粒pHTL傳10代后質(zhì)粒仍有98.5%的存在。

對比試驗：接種重組工程菌B8- pHT315gfp（構(gòu)建過程中g(shù)fp插入pHT315的多克隆位點處，構(gòu)建過程與pHTLgfp相同，省略構(gòu)建過程）于含有紅霉素的LB液體培養(yǎng)基中培養(yǎng)活化轉(zhuǎn)接至LB液體培養(yǎng)基中搖瓶培養(yǎng) 12 h 后，以1%的接種量轉(zhuǎn)入新的LB液體性培養(yǎng)基中（不含紅霉素），開始連續(xù)搖瓶培養(yǎng)試驗。每12 h按1%接種量轉(zhuǎn)接一次為一代工轉(zhuǎn)接10代。 每一代取樣1 mL進(jìn)行平板稀釋涂布檢測質(zhì)粒的缺失程度。檢測時，取菌液用無菌水逐級稀釋，取三個合適的梯度，分別涂在含有和不含有紅霉素的固體LB培養(yǎng)基平板上，每個處理三個重復(fù)。 倒置在 37℃培養(yǎng)箱中培養(yǎng)24h，計算各皿中的菌落數(shù)。計算出質(zhì)粒的穩(wěn)定率（穩(wěn)定率=含有抗生素平板的菌落數(shù)/不含有抗生素平板的菌落數(shù)）。結(jié)果表明在無抗生素選擇壓力下質(zhì)粒pHT315傳10代后質(zhì)粒只有48.9%的存在。

實施例2：

（1）納豆芽孢桿菌基因組提取

無菌條件下，挑取納豆芽孢桿菌單菌落，接種于LB培養(yǎng)基中，震蕩培養(yǎng)過夜，轉(zhuǎn)接到20 mL的LB培養(yǎng)基中繼續(xù)震蕩培養(yǎng)過夜；將菌液收集在離心管中，離心收集所有菌體，用TE洗一次；加入200 μL TE緩沖液震蕩混勻；加入200 μL 2%的SDS緩沖液混勻；加入400 μL異硫氰酸胍已酸鈉混合液；12000 rpm離心5 min，將上清轉(zhuǎn)移到一個新的離心管中，加入600 μL異丙醇混勻；12000 rpm離心10 min去上清，加入500 μL 70%乙醇混勻；12000 rpm離心10 min，倒去上清，室溫晾干至無乙醇?xì)馕?；加?0 μL滅菌的去離子水，加入5 μLRNA酶溶液，37 ℃孵育30 min，用1%的瓊脂糖凝膠進(jìn)行電泳，檢測提取的的基因組DNA。

（2）融合片段的擴(kuò)增

分別以質(zhì)粒pHTL為模板用引物對xyls-5/xyls-3（SEQ ID NO:18/ SEQ ID NO:19）擴(kuò)增1167bp的木糖啟動子部分序列PxylRstuI；以納豆芽孢桿菌基因組為模板，用引物對apre-5/apre-3（SEQ ID NO:20/ SEQ ID NO:21）擴(kuò)增得到納豆芽孢桿菌中96bp的信號肽Apre表達(dá)區(qū)片段；回收兩個PCR產(chǎn)物，不添加引物，利用PrimeSTAR Max DNA Polymerase進(jìn)行互補(bǔ)延伸，形成融合片段的中間產(chǎn)物，以此為模板，用引物對xyls-5/apre-3進(jìn)行融合PCR的擴(kuò)增，獲得木糖啟動子部分序列PxylRstuI和納豆芽孢桿菌信號肽表達(dá)區(qū)的融合片段PxylRstuI-Apre，大小為1263bp，經(jīng)測序驗證正確性。

（3） 分泌型穿梭表達(dá)載體pHTZ的構(gòu)建

融合片段PxylRstuI-Apre用限制性內(nèi)切酶Stu I/BamH I雙酶切并進(jìn)行膠回收后，連接到經(jīng)同種酶酶切后的載體pHTL上，獲得重組質(zhì)粒pHTZ。該質(zhì)粒含有PxylR序列、Apre序列、多克隆位點區(qū)、T7終止子，物理圖譜如圖10。重組質(zhì)粒經(jīng)多種限制性內(nèi)切酶單酶切、PCR及測序鑒定(SEQ ID NO:6/ SEQ ID NO:7)，證明構(gòu)建正確圖11，序列見（SEQ ID NO:17）。

（4） 高效分泌型穿梭表達(dá)載體的效率評估

質(zhì)粒pHTL-gus經(jīng)限制性內(nèi)切酶Xho I/Hind III雙酶切后得到大小為1812bp的gus基因；質(zhì)粒pHTL-tgl1經(jīng)限制性內(nèi)切酶Sal I/Hind III雙酶切后得到大小為738bp的tgl1基因；以質(zhì)粒pHTL-lipa經(jīng)限制性內(nèi)切酶Sal I/Hind III雙酶切后得到大小為1842 bp的lipa基因?；厥蘸蟮幕蚱螌?dǎo)入pHTZ的相應(yīng)酶切位點間，轉(zhuǎn)化E. coli JM109，得到重組質(zhì)粒pHTZ-gus，pHTZ-tgl1，pHTZ-lipa，經(jīng)酶切測序鑒定正確（圖12），證明構(gòu)建正確，序列見（SEQ ID NO:23、SEQ ID NO:24、SEQ ID NO:25）。再轉(zhuǎn)入E. coli SCS110去甲基化后，采用熱擊轉(zhuǎn)化的方法將重組質(zhì)粒導(dǎo)入B.subtilis WB800感受態(tài)細(xì)胞中，獲得B8-pHTZgus，B8-pHTZtgl1和B8-pHTZlipa工程菌。

工程菌B8-pHTZgus中GUS表達(dá)檢測：挑取工程菌單菌落，接種于5 mL含有紅霉素(10 μg /mL )LB液體培養(yǎng)基中，活化后轉(zhuǎn)接新的液體培養(yǎng)基中震蕩培養(yǎng)至菌體OD為0.5時，加入終濃度為2%的D-Xylose，誘導(dǎo)24 h后收獲。取1 mL培養(yǎng)基，離心收集上清。加入450 μLGUS基液和50 μL的X-Gluc母液，混勻。37 ℃保溫，肉眼觀察染色結(jié)果（圖13）。證明分泌型穿梭表達(dá)載體pHTZ能成功的高效表達(dá)GUS蛋白。

工程菌B8-pHTZtgl1中谷氨酰胺轉(zhuǎn)氨酶的表達(dá)：將活化的工程菌轉(zhuǎn)接LB液體培養(yǎng)基中震蕩培養(yǎng)，當(dāng)菌體OD為0.5時，加入終濃度為2％的D-Xylose，誘導(dǎo)24h后離心收獲上清，與BSA(1mg/mL)混合后于70℃溫育3h，利用SDS-PAGE電泳檢測tgl1蛋白含量的表達(dá)情況。SDS-PAGE電泳檢測方法參見分子克隆實驗指南[J.薩姆布魯克。分子克隆實驗指南(第二版，金冬雁等).北京:科學(xué)出版社,1992]。結(jié)果證明工程菌表達(dá)的Tgl1催化BSA發(fā)生交聯(lián)(圖13)，而B.subtilis WB800菌液與BSA混合后未檢測出BSA的聚合條帶，說明分泌型穿梭表達(dá)載體pHTZ可以成功地表達(dá)Tgl1蛋白。

工程菌B8-pHTZlipa中脂肪酶的表達(dá)：建立對硝基苯酚（p-NP）的濃度標(biāo)準(zhǔn)曲線。將活化的工程菌B8-pHTZlipa轉(zhuǎn)接LB液體培養(yǎng)基中震蕩培養(yǎng)，當(dāng)菌體OD為0.5時，加入終濃度為2%的D-Xylose，誘導(dǎo)24h后收獲。離心收獲上清液進(jìn)行酶活測定。同時，收獲以同樣接種培養(yǎng)同時期收獲但不加D-Xylose的工程菌B8-pHTZlipa和以同樣接種培養(yǎng)同時期收獲的WB800菌體，作為脂肪酶表達(dá)的參照。根據(jù)建立p-NP濃度與410 nm下吸光值的標(biāo)準(zhǔn)曲線，計算破碎液中的p-NP量，根據(jù)脂肪酶活力定義計算收獲的上清中脂肪酶的酶活（U/mg），作為對照的WB800幾乎沒有脂肪酶活性（0.25±0.07 U/mg），同時，無木糖誘導(dǎo)的工程菌B8-pHTZlipa的菌體酶活力為0.41±0.09 U/mg，誘導(dǎo)后的脂肪酶活力為181.2±1.75 U/mg，提高了441倍，說明載體pHTZ成功并高效表達(dá)了LipA蛋白。

對比試驗：建立對硝基苯酚（p-NP）的濃度標(biāo)準(zhǔn)曲線。將活化的工程菌B8-pHTLlipa（構(gòu)建過程lipa的插入位點與pHTZlipa相同，省略構(gòu)建過程）轉(zhuǎn)接LB液體培養(yǎng)基中震蕩培養(yǎng)，當(dāng)菌體OD為0.5時，加入終濃度為2%的D-Xylose，誘導(dǎo)24h后收獲。離心收獲上清液進(jìn)行酶活測定。同時，收獲以同樣接種培養(yǎng)同時期收獲但不加D-Xylose的工程菌B8-pHTLlipa和以同樣接種培養(yǎng)同時期收獲的WB800菌體，作為脂肪酶表達(dá)的參照。根據(jù)建立p-NP濃度與410 nm下吸光值的標(biāo)準(zhǔn)曲線，計算破碎液中的p-NP量，根據(jù)脂肪酶活力定義計算收獲的上清中脂肪酶的酶活（U/mg），作為對照的WB800無論有無木糖誘導(dǎo)都只有微弱的脂肪酶活性（0.22±0.08 U/mg），無木糖誘導(dǎo)的工程菌B8-pHTLlipa同樣也只有微弱的脂肪酶活性（0.39±0.11 U/mg），證明在沒有木糖誘導(dǎo)的時候，木糖啟動子幾乎沒有啟動目的基因的轉(zhuǎn)錄，而當(dāng)有木糖誘導(dǎo)時，木糖啟動子則會表現(xiàn)出相當(dāng)強(qiáng)大的啟動目的基因轉(zhuǎn)錄的功能，上述事實表明pHTZ是一個典型的誘導(dǎo)型表達(dá)載體。

SEQUENCE LISTING

<110> 河北農(nóng)業(yè)大學(xué)

<120> 一種高效穿梭表達(dá)載體及其構(gòu)建方法與應(yīng)用

<130> 1111

<160> 24

<170> Patent In version 3.5

<210> 1

<211> 8170

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 1

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca catggctagc 7380

atgactggtg gacagcaaat gggtcgggat ccgaattcga gctccgtcga caagcttgcg 7440

gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa agcccgaaag 7500

gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct 7560

aaacgggtct tgaggggttt tttgcccaat tcactggccg tcgttttaca acgtcgtgac 7620

tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc 7680

tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat 7740

ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc 7800

atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac 7860

ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga 7920

caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa 7980

cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata 8040

atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt 8100

ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg 8160

cttcaataat 8170

<210> 2

<211> 24

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 2

ggcccgggaa atactcctag aata 24

<210> 3

<211> 36

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 3

ccaccagtca tgctagccat gtgatttccc ccttaa 36

<210> 4

<211> 36

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 4

ttaaggggga aatcacatgg ctagcatgac tggtgg 36

<210> 5

<211> 28

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 5

aaatcccggg caaaaaaccc ctcaagac 28

<210> 6

<211> 18

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 6

caggaaacag ctatgacc 18

<210> 7

<211> 18

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 7

tgtaaaacga cggccagt 18

<210> 8

<211> 24

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 8

ccggaattcg atggtagatc tgac 24

<210> 9

<211> 24

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 9

cccaagcttt cacacgtggt ggtg 24

<210> 10

<211> 32

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 10

cggaagctta tgttacgtcc tgtagaaacc cc 32

<210> 11

<211> 30

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 11

taactcgagt cattgtttgc ctccctgctg 30

<210> 12

<211> 24

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 12

aacgtcgaca tgggcatctt tagc 24

<210> 13

<211> 24

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 13

ggaaagcttt taggccaaca ccac 24

<210> 14

<211> 8914

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 14

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca catggctagc 7380

atgactggtg gacagcaaat gggtcgggat ccgaattcga tggtagatct gactagtaaa 7440

ggagaagaac ttttcactgg agttgtccca attcttgttg aattagatgg tgatgttaat 7500

gggcacaaat tttctgtcag tggagagggt gaaggtgatg caacatacgg aaaacttacc 7560

cttaaattta tttgcactac tggaaaacta cctgttccgt ggccaacact tgtcactact 7620

ttctcttatg gtgttcaatg cttttcaaga tacccagatc atatgaagcg gcacgacttc 7680

ttcaagagcg ccatgcctga gggatacgtg caggagagga ccatcttctt caaggacgac 7740

gggaactaca agacacgtgc tgaagtcaag tttgagggag acaccctcgt caacaggatc 7800

gagcttaagg gaatcgattt caaggaggac ggaaacatcc tcggccacaa gttggaatac 7860

aactacaact cccacaacgt atacatcatg gccgacaagc aaaagaacgg catcaaagcc 7920

aacttcaaga cccgccacaa catcgaagac ggcggcgtgc aactcgctga tcattatcaa 7980

caaaatactc caattggcga tggccctgtc cttttaccag acaaccatta cctgtccaca 8040

caatctgccc tttcgaaaga tcccaacgaa aagagagacc acatggtcct tcttgagttt 8100

gtaacagctg ctgggattac acatggcatg gatgaactat acaaagctag ccaccaccac 8160

caccaccacg tgtgaaagct tgcggccgca ctcgagcacc accaccacca ccactgagat 8220

ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa 8280

ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgcc caattcactg 8340

gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt 8400

gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct 8460

tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt tctccttacg 8520

catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg ctctgatgcc 8580

gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt 8640

ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag 8700

aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt 8760

ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga 8820

aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc 8880

atgagacaat aaccctgata aatgcttcaa taat 8914

<210> 15

<211> 9973

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 15

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca catggctagc 7380

atgactggtg gacagcaaat gggtcgggat ccgaattcga gctccgtcga caagcttatg 7440

ttacgtcctg tagaaacccc aacccgtgaa atcaaaaaac tcgacggcct gtgggcattc 7500

agtctggatc gcgaaaactg tggaattgat cagcgttggt gggaaagcgc gttacaagaa 7560

agccgggcaa ttgctgtgcc aggcagtttt aacgatcagt tcgccgatgc agatattcgt 7620

aattatgcgg gcaacgtctg gtatcagcgc gaagtcttta taccgaaagg ttgggcaggc 7680

cagcgtatcg tgctgcgttt cgatgcggtc actcattacg gcaaagtgtg ggtcaataat 7740

caggaagtga tggagcatca gggcggctat acgccatttg aagccgatgt cacgccgtat 7800

gttattgccg ggaaaagtgt acgtatcacc gtttgtgtga acaacgaact gaactggcag 7860

actatcccgc cgggaatggt gattaccgac gaaaacggca agaaaaagca gtcttacttc 7920

catgatttct ttaactatgc cggaatccat cgcagcgtaa tgctctacac cacgccgaac 7980

acctgggtgg acgatatcac cgtggtgacg catgtcgcgc aagactgtaa ccacgcgtct 8040

gttgactggc aggtggtggc caatggtgat gtcagcgttg aactgcgtga tgcggatcaa 8100

caggtggttg caactggaca aggcactagc gggactttgc aagtggtgaa tccgcacctc 8160

tggcaaccgg gtgaaggtta tctctatgaa ctgtgcgtca cagccaaaag ccagacagag 8220

tgtgatatct acccgcttcg cgtcggcatc cggtcagtgg cagtgaaggg cgaacagttc 8280

ctgattaacc acaaaccgtt ctactttact ggctttggtc gtcatgaaga tgcggacttg 8340

cgtggcaaag gattcgataa cgtgctgatg gtgcacgacc acgcattaat ggactggatt 8400

ggggccaact cctaccgtac ctcgcattac ccttacgctg aagagatgct cgactgggca 8460

gatgaacatg gcatcgtggt gattgatgaa actgctgctg tcggctttaa cctctcttta 8520

ggcattggtt tcgaagcggg caacaagccg aaagaactgt acagcgaaga ggcagtcaac 8580

ggggaaactc agcaagcgca cttacaggcg attaaagagc tgatagcgcg tgacaaaaac 8640

cacccaagcg tggtgatgtg gagtattgcc aacgaaccgg atacccgtcc gcaaggtgca 8700

cgggaatatt tcgcgccact ggcggaagca acgcgtaaac tcgacccgac gcgtccgatc 8760

acctgcgtca atgtaatgtt ctgcgacgct cacaccgata ccatcagcga tctctttgat 8820

gtgctgtgcc tgaaccgtta ttacggatgg tatgtccaaa gcggcgattt ggaaacggca 8880

gagaaggtac tggaaaaaga acttctggcc tggcaggaga aactgcatca gccgattatc 8940

atcaccgaat acggcgtgga tacgttagcc gggctgcact caatgtacac cgacatgtgg 9000

agtgaagagt atcagtgtgc atggctggat atgtatcacc gcgtctttga tcgcgtcagc 9060

gccgtcgtcg gtgaacaggt atggaatttc gccgattttg cgacctcgca aggcatattg 9120

cgcgttggcg gtaacaagaa agggatcttc actcgcgacc gcaaaccgaa gtcggcggct 9180

tttctgctgc aaaaacgctg gactggcatg aacttcggtg aaaaaccgca gcagggaggc 9240

aaacaatgac tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga 9300

aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc 9360

tctaaacggg tcttgagggg ttttttgccc aattcactgg ccgtcgtttt acaacgtcgt 9420

gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc 9480

agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg 9540

aatggcgaat ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac 9600

cgcatatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga 9660

cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac 9720

agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg 9780

aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata 9840

ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt 9900

tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa 9960

atgcttcaat aat 9973

<210> 16

<211> 10015

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 16

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca catggctagc 7380

atgactggtg gacagcaaat gggtcgggat ccgaattcga gctccgtcga catgggcatc 7440

tttagctata aggatttgga cgaaaacgcg tcgaaagcgc tgttttccga cgccttggcc 7500

atctccacct acgcttacca caatatcgat aacggcttcg acgaaggcta ccaccagacc 7560

ggtttcggtc ttggcctgcc gctgacgctg atcaccgcgc tgatcggcag cacccaatcg 7620

cagggcggcc tgccccgcat tccctggaac cccgactccg aacaggccgc gcaggagacg 7680

gtgaacaatg ccggctggtc ggtcatcagc gccgcgcagc tgggttacgc cggcaaaacc 7740

gatgcacgcg gcacctatta cggcgagacc gccggttaca ccaccgcgca ggccgaggtg 7800

ctgggcaaat atgacagcga aggcaatctc accgccattg gtatctcatt tcgcggtacc 7860

agcggcccgc gcgagtcgct gatcggcgat accatcggcg atgtgattaa cgatctgctg 7920

gccggtttcg ggccgaaagg ctacgctgac ggctacacgc tgaacgcctt cggcaatctg 7980

ctgggcgacg tggcgaaatt cgcgcaggcg cacgggctga gcggcgagga cgtagtggtc 8040

agcggccaca gcctcggcgg gctggcggtc aacagcatgg cggcgcagag cgacgccaac 8100

tggggcggct tctacgcgca gtccaactat gtcgccttcg cctcgccgac ccagtacgaa 8160

gccggcggca aggtgatcaa catcggctac gagaacgacc cggtgttccg cgcgctcgac 8220

ggcacctcgc taaccctgcc gtcactgggc gtacacgatg cgccgcacgc ctccgccacc 8280

aacaatatcg tcaacttcaa cgaccactac gcgtcggacg cctggaacct gctgccgttt 8340

tccattctca acattccgac ctggctgtcg cacctgccgt tcttctatca ggacgggctg 8400

atgcgggtgc tgaactccga gttttattcg ctgaccgaca aggactcgac catcatcgtc 8460

tccaacctgt cgaacgtgac gcgcggcaat acctgggtgg aagacctgaa ccgcaacgcg 8520

gaaacgcaca gcggaccgac gtttatcatc ggcagcgacg gcaatgattt gatcaagggc 8580

ggcaaaggca acgactatct cgagggccgc gacggcgacg atatcttccg cgacgccggc 8640

ggctataacc tgatcgccgg cggcaaaggc cacaatatct tcgataccca acaggcgttg 8700

aaaaacaccg aggtcgccta cgacggcaat acgctttacc tgcgcgacgc caaaggcggt 8760

attacgctgg cagacgacat cagcaccctg cgcagcaaag aaacctcctg gctgattttc 8820

agcaaagagg tggatcatca ggtgaccgct gcgggattga aatcggactc gggcctcaaa 8880

gcctatgccg ccgccaccac cggcggcgac ggcgatgacg tcctgcaggc tcgcagccac 8940

gacgcctggc tgttcggcaa cgccggcaac gacacgctga tcggccatgc cggcggcaac 9000

ctgaccttcg tcggcggcag cggcgatgac atcctgaagg gcgccggcaa cggtaatacc 9060

ttcctgttca gcggcgattt cggccgcgac cagctgtatg gtttcaacgc caccgataaa 9120

ctggtgttta tcggtaccga aggcgccagc gggaatatcc gcgactatgc cacacagcaa 9180

aacgacgatc tggtgctggc cttcggccac ggccaggtca cgctgatcgg cgtctcgctc 9240

gatcacttca acaccgatcg ggtggtgttg gcctaaaagc ttgcggccgc actcgagcac 9300

caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct 9360

gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg 9420

ggttttttgc ccaattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc 9480

gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa 9540

gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg 9600

atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc 9660

agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct 9720

gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc 9780

tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag 9840

ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg 9900

tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata 9960

cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataat 10015

<210> 17

<211> 8230

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 17

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca cgtgagaagc 7380

aaaaaattgt ggatcagctt gttgtttgcg ttaacgttaa tctttacgat ggcgttcagc 7440

aacatgtctg cgcaggctgc cggcaaggat ccgaattcga gctccgtcga caagcttgcg 7500

gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa agcccgaaag 7560

gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct 7620

aaacgggtct tgaggggttt tttgcccaat tcactggccg tcgttttaca acgtcgtgac 7680

tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc 7740

tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat 7800

ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc 7860

atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac 7920

ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga 7980

caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa 8040

cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata 8100

atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt 8160

ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg 8220

cttcaataat 8230

<210> 18

<211> 25

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 18

ggaggcctat tcctaaatag aatcc 25

<210> 19

<211> 33

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 19

ccacaatttt ttgcttctca cgtgatttcc ccc 33

<210> 20

<211> 33

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 20

gggggaaatc acgtgagaag caaaaaattg tgg 33

<210> 21

<211> 30

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 21

ttaattaatt ggatccttgc cggcagcctg 30

<210> 22

<211> 10033

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 22

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca cgtgagaagc 7380

aaaaaattgt ggatcagctt gttgtttgcg ttaacgttaa tctttacgat ggcgttcagc 7440

aacatgtctg cgcaggctgc cggcaaggat ccgaattcga gctccgtcga caagcttatg 7500

ttacgtcctg tagaaacccc aacccgtgaa atcaaaaaac tcgacggcct gtgggcattc 7560

agtctggatc gcgaaaactg tggaattgat cagcgttggt gggaaagcgc gttacaagaa 7620

agccgggcaa ttgctgtgcc aggcagtttt aacgatcagt tcgccgatgc agatattcgt 7680

aattatgcgg gcaacgtctg gtatcagcgc gaagtcttta taccgaaagg ttgggcaggc 7740

cagcgtatcg tgctgcgttt cgatgcggtc actcattacg gcaaagtgtg ggtcaataat 7800

caggaagtga tggagcatca gggcggctat acgccatttg aagccgatgt cacgccgtat 7860

gttattgccg ggaaaagtgt acgtatcacc gtttgtgtga acaacgaact gaactggcag 7920

actatcccgc cgggaatggt gattaccgac gaaaacggca agaaaaagca gtcttacttc 7980

catgatttct ttaactatgc cggaatccat cgcagcgtaa tgctctacac cacgccgaac 8040

acctgggtgg acgatatcac cgtggtgacg catgtcgcgc aagactgtaa ccacgcgtct 8100

gttgactggc aggtggtggc caatggtgat gtcagcgttg aactgcgtga tgcggatcaa 8160

caggtggttg caactggaca aggcactagc gggactttgc aagtggtgaa tccgcacctc 8220

tggcaaccgg gtgaaggtta tctctatgaa ctgtgcgtca cagccaaaag ccagacagag 8280

tgtgatatct acccgcttcg cgtcggcatc cggtcagtgg cagtgaaggg cgaacagttc 8340

ctgattaacc acaaaccgtt ctactttact ggctttggtc gtcatgaaga tgcggacttg 8400

cgtggcaaag gattcgataa cgtgctgatg gtgcacgacc acgcattaat ggactggatt 8460

ggggccaact cctaccgtac ctcgcattac ccttacgctg aagagatgct cgactgggca 8520

gatgaacatg gcatcgtggt gattgatgaa actgctgctg tcggctttaa cctctcttta 8580

ggcattggtt tcgaagcggg caacaagccg aaagaactgt acagcgaaga ggcagtcaac 8640

ggggaaactc agcaagcgca cttacaggcg attaaagagc tgatagcgcg tgacaaaaac 8700

cacccaagcg tggtgatgtg gagtattgcc aacgaaccgg atacccgtcc gcaaggtgca 8760

cgggaatatt tcgcgccact ggcggaagca acgcgtaaac tcgacccgac gcgtccgatc 8820

acctgcgtca atgtaatgtt ctgcgacgct cacaccgata ccatcagcga tctctttgat 8880

gtgctgtgcc tgaaccgtta ttacggatgg tatgtccaaa gcggcgattt ggaaacggca 8940

gagaaggtac tggaaaaaga acttctggcc tggcaggaga aactgcatca gccgattatc 9000

atcaccgaat acggcgtgga tacgttagcc gggctgcact caatgtacac cgacatgtgg 9060

agtgaagagt atcagtgtgc atggctggat atgtatcacc gcgtctttga tcgcgtcagc 9120

gccgtcgtcg gtgaacaggt atggaatttc gccgattttg cgacctcgca aggcatattg 9180

cgcgttggcg gtaacaagaa agggatcttc actcgcgacc gcaaaccgaa gtcggcggct 9240

tttctgctgc aaaaacgctg gactggcatg aacttcggtg aaaaaccgca gcagggaggc 9300

aaacaatgac tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga 9360

aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc 9420

tctaaacggg tcttgagggg ttttttgccc aattcactgg ccgtcgtttt acaacgtcgt 9480

gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc 9540

agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg 9600

aatggcgaat ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac 9660

cgcatatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga 9720

cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac 9780

agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg 9840

aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata 9900

ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt 9960

tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa 10020

atgcttcaat aat 10033

<210> 23

<211> 8968

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 23

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca cgtgagaagc 7380

aaaaaattgt ggatcagctt gttgtttgcg ttaacgttaa tctttacgat ggcgttcagc 7440

aacatgtctg cgcaggctgc cggcaaggat ccgaattcga gctccgtcga catgattatt 7500

gtatcaggac aattgctccg tccccaggat attgaaaatt ggcagattga tcaagatctg 7560

aatccgctct taaaagagat gattgagacg cctgttcagt ttgattatca ttcaattgct 7620

gaactgatgt ttgagcttaa actgcggatg aatattgtag cagcggcaaa gacgctgcac 7680

aaaagcgggg cgaagtttgc cactttttta aaaacatacg ggaatacaac gtattggagg 7740

gtttcaccgg agggcgcctt ggagctgaaa tacagaatgc cgccttcaaa agcgattcgg 7800

gacattgcag agaacggccc gttttatgcg tttgaatgcg caaccgcaat cgttatcatt 7860

tattacttgg ccttaatcga tacaatcgga gaagataaat tcaatgccag ctttgacaga 7920

attattttat atgactggca ttatgagaaa ttgccgatat atacggaaac aggacaccac 7980

tttttccttg gagattgttt gtattttaag aatcctgaat ttgatccgca aaaggcgcaa 8040

tggagaggcg aaaatgtgat actactgggg gaagataaat attttgccca tggtcttgga 8100

atcttaaacg gaaagcaaat tattgataag ctgaattctt ttaggaaaaa aggagcctta 8160

cagtcagcct accttctgtc tcaggcgacc agactggatg ttccgtctct tttccgcatc 8220

gtccgctaaa agcttgcggc cgcactcgag caccaccacc accaccactg agatccggct 8280

gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca 8340

taaccccttg gggcctctaa acgggtcttg aggggttttt tgcccaattc actggccgtc 8400

gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca 8460

catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa 8520

cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct tacgcatctg 8580

tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag 8640

ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc 8700

ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt 8760

tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag 8820

gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg 8880

cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga 8940

caataaccct gataaatgct tcaataat 8968

<210> 24

<211> 10075

<212> DNA

<213> Artificial Sequence

<220>

<223> Artificial Sequence

<400> 24

ccatcctcca aagttggaga gtgagtttta tgtcgcaaat attaatgttt ctggtgaacc 60

ttatcaaatt ttcgttgatt taatagaaac atagcggtaa aattagcagt aacttaatag 120

aacggaaatg aaaaaagcca ctctcatatg ctattggcta ccaaccttta gcgagaatga 180

cttaatcctg tacagccata caggacttcg acttataaga ggcgccaacc tcaaataagt 240

tatttgcctt gttttcgcga acaaggctta ttagatacac ctattgtacc gttactctac 300

gaatatttca actagtaatt actagcattg tcatatacat aataaaacgg atataaaagg 360

gcgttttcta tacctagaag tcttgtaaat gtacagggcg tttagatata gagaacgccc 420

tttttgtgtt ccgttccagt ggaagctacc actttaaaaa gatggtctag tgtagccaat 480

gcaggagagt acactcggat atcagttgtc gttgcattca actgtctgac gtaagcgagg 540

taaaggacac aagccttgca taaaacaagc ctacgggatg taaatcctaa taatgatgat 600

aaccaagacg ttagcggcaa aaagtgttgg gggttcaaaa taagacatga ttgtgcgact 660

ggagttaaac agttactcgt aagcggcgat catgacactg attcacggct attcttgtac 720

aagctagctt tattacaagg atatgcgggt tatatagcga atcacccgaa agggaacggt 780

gttgggcgtg agaaacgcac cgtacggcgc aatacaatgc caataagcta tatacggacg 840

gtatagtagt tttgtaagct ataaccgttt gtcgtcaatg caaccaatct caattcgaga 900

cctcggcatc taagccagta cgaatgagtg ggcgttttaa cctcgtaaat tttcaacagg 960

ggttactatg cccaaaacta cattcagatt tcctaacaaa ctcgccagta tgaaaacctt 1020

aagaccttaa agtcaaggga tttgaaggat tttaacctcg attagcaaaa aatgtagagt 1080

actgaagcaa ctaccattaa ctaagatagt gggggattga ggaagaatcc agagctgttt 1140

aaatcaagtg aaagacaaga tgaaattaaa agaatagtga aagatagggg agtggttctc 1200

tatgagaaag gaaatggcta gagaacaaag gcagcggttt attgatctat tgttagactt 1260

tatggtaaag aatcctcatt tatttgttaa tggtacagag gatgaaagta ataatgttgt 1320

tacaaaatgt aatagtgata ttaaagaggt tgcggagtca tatttaactc ttttatagtg 1380

agagggttaa aactaattaa tatgtattaa ggcccaatgt tggaattatt gtatttcact 1440

aggcaaccta cttactaaaa gtaagattat ccattagtgg atgttataat attgggtttt 1500

ttaacacaat aatcatcgcc tttcggtgtc gtttgataga aaagtaacca ttagcgatga 1560

aaaagtcaat ataaaaagcc atccgtaaaa aacggatggc ttaccgtaca taggatcgtt 1620

ggtagggcgg cgtatcctac atctctggta acttacctag ccaatcaaat gcttgagaac 1680

ggcggttaga taagcgcgtg gggaaccttt cccacctcaa agatcctata tcattattat 1740

gttactttct acaggtagta taccatgttc ttatatttta gtaaactccc cgttagctta 1800

acaggtcttt gtaagcaatt aaacgtccac tattcaatcg tctttggatt ttcgcaggac 1860

cgttttttag atcgaacata gttgataaga acaaataacc gcttgggtcc aactttatag 1920

caattagtat atggtcattt aaaatcttta ccaattcaac gctattaggt tctttaggat 1980

tttgcccgac atagtcgggg tgttcaacga tatcttttat gtgcgatgaa tatttttcat 2040

aaataccagg atgttgtttc tttacgtgct ttataaatcc gggaaacatt tttacatcgt 2100

tagaagtgca agtcaagtta tatgtatcta taatgatttg tggaagtttt gccacaacag 2160

ttggtttatt tacaatcttt tttttattag ccgtcaaatt tctccctcat ctcgtctctt 2220

tatatcttta ttttatcata aaggagtatt tgaaccgtcg cgcgggacag gtttatgata 2280

gggatatttt attgaataat tgatggtata agggactttc atgcttggaa agtggggatt 2340

atgaattaga tgcttgtcca caatatgttc caatgtaatt aaaatttatg ttcccacctt 2400

gaccaaacat cacgtccata cttaaatcgt ccctccttta ataggtaaaa tattaattta 2460

ccttaataaa aaaataatgg ataatagtat tcgtctgaat ttatataatc agggggaact 2520

attgatgctg gggatactat ttacagcggc gccatctact gatgtcgtaa aggatttgca 2580

agataaagtt atatcattgc aggatcatga ggtagcgttt ttgaacacca cgatatctaa 2640

tatgttgatc ccccttagaa gcaaacttaa gagtgtgttg atagtgcagt atcttaaaat 2700

tttgtgtata ataggaattg aagttaaatt agatgctaaa aatttgtaat taagaaggag 2760

ggattcgtca tgttggtatt ccaaatgcgt aatgtagata aaacatctac tgttttgaaa 2820

cagactaaaa acagtgatta cgcagataaa taaatacgtt agattaattc ctaccagtga 2880

ctaatcttat gactttttaa acagataact aaaattacaa acaaatcgtt taacttctgt 2940

atttatttac agatgtaatc acttcaggag taattacatg aacaaaaata taaaatattc 3000

tcaaaacttt ttaacgagtg aaaaagtact caaccaaata ataaaacaat tgaatttaaa 3060

agaaaccgat accgtttacg aaattggaac aggtaaaggg catttaacga cgaaactggc 3120

taaaataagt aaacaggtaa cgtctattga attagacagt catctattca acttatcgtc 3180

agaaaaatta aaactgaaca ttcgtgtcac tttaattcac caagatattc tacagtttca 3240

attccctaac aaacagaggt ataaaattgt tgggagtatt ccttaccatt taagcacaca 3300

aattattaaa aaagtggttt ttgaaagcca tgcgtctgac atctatctga ttgttgaaga 3360

aggattctac aagcgtacct tggatattca ccgaacacta gggttgctct tgcacactca 3420

agtctcgatt cagcaattgc ttaagctgcc agcggaatgc tttcatccta aaccaaaagt 3480

aaacagtgtc ttaataaaac ttacccgcca taccacagat gttccagata aatattggaa 3540

gctatatacg tactttgttt caaaatgggt caatcgagaa tatcgtcaac tgtttactaa 3600

aaatcagttt catcaagcaa tgaaacacgc caaagtaaac aatttaagta ccattactta 3660

tgagcaagta ttgtctattt ttaatagtta tctattattt aacgggagga aataattcta 3720

tgagtcgctt ttttaaattt ggaaagttac acgttactaa agggaatgga gataaattat 3780

tagatatact actgacagct tccaagaagc taaagaggtc cctagcgcct acggggaatt 3840

tgtatcgcga tgggtacatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 3900

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 3960

gaaagtaaaa gatgctgaag atctgttggg tgcacgagtg ggttacatcg aactggatct 4020

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 4080

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 4140

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 4200

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 4260

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 4320

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 4380

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 4440

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 4500

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 4560

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 4620

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 4680

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 4740

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4800

gatctaggtg aagatccttt ttgataatct tcatgaccaa aatcccttaa cgtgagtttt 4860

cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt 4920

ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 4980

tgccgatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5040

accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5100

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5160

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5220

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5280

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5340

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5400

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5460

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5520

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5580

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 5640

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 5700

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 5760

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 5820

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 5880

aggaaacagc tatgaccatg attacgccaa gctgggaaat actcctagaa taaaaaaact 5940

catctttaaa gatgagctgt ccattccata aaaaattaca ttgtaatcat gtccagaaaa 6000

tgatcaatca caatggagga cattcctaat gccggtgcat tctgtcctaa ggaagatggc 6060

aataattcat agctattgcc taattgggaa taaacccttg atgatacttc acttctcatt 6120

gaatttaaaa ccataggatg cgattcaatt atgctatttc ttaaaattac ggcttgtggg 6180

ttgaaagtat ttagaatatt ggtaaggcct attcctaaat agaatccaaa attttgtaat 6240

gcatttaagg ttccgatatc attcagatgg gcgaggttta tgatatcttg ataggacagt 6300

tttttctctt tggtctgaag agattttaat aaagccttct ctgaagcata caattcccag 6360

catcctcggt ttccgcaact gcatttagga ccattaaagt ctattgtcat atgtcccatt 6420

tctccagaga agccgcttac tcctctatat aaatgattgt tgataataac accgatccct 6480

attcctgtgc tgatacttac gtaaataatg ttatcgtgat tttttgcagc tccaaatact 6540

ttttctccat atgcgccagc atttgcctca ttttcaataa aaacaggcac attgtacttc 6600

tcttgtatcg aagattttaa gtcaatatct ctccagttgg agttcggagt gaaaacaatt 6660

ttttgatctt tatcaatgag tccaggcacg caaataccta taccaataag cccgtacgga 6720

gattggggca tttgcgtaat aaagtgatga atcatatcaa tcaaaatgtc tttcgttatt 6780

tctggagaat tggattccaa atggcggtat tgatcaagaa cgattgttcc ttcaaggtct 6840

gttaaaatgc cattaatata atccacacca acatctattc caacggagta tcctgccttt 6900

ttattaaaaa caagcatgac aggtcttctt ccgccacttg attgtccttg acctatttca 6960

aataccatac tttctttcat taacgtgttt acctgtgatg agacagttga tttatttaat 7020

ccagtcattt cagataattt tgctcttgaa ataggtgaat ttttaaggat ttcttttaat 7080

aataactttt gatttacttt tttgacaaag gtttgatcag cgatatccac ttcatccact 7140

ccatttgttt aatctttaaa ttaagtatca acatagtaca tagcgaatct tccctttatt 7200

atatctaatg tgttcataaa aaactaaaaa aaatattgaa aatactgacg aggttatata 7260

agatgaaaat aagttagttt gtttaaacaa caaactaata ggtgatgtac ttactatatg 7320

aaataaaatg catctgtatt tgaatgaatt tatttttaag ggggaaatca cgtgagaagc 7380

aaaaaattgt ggatcagctt gttgtttgcg ttaacgttaa tctttacgat ggcgttcagc 7440

aacatgtctg cgcaggctgc cggcaaggat ccgaattcga gctccgtcga catgggcatc 7500

tttagctata aggatctgga cgaaaatgcg tcgaaggcgc tgttttccga cgccttggcc 7560

atctctacct acgcttacca taatatcgat aacggcttcg atgaaggcta tcaccagacc 7620

ggtttcggcc tcggtctgcc actgacgctg gtcactgcgc tgatcggcag tacccagtcg 7680

cagggtggcc tgcctggcct cccttggaat cccgactccg aacaggccgc gcaggaggcg 7740

gtaaacaatg ccggctggtc ggtgatcagc gccgcgcagc tcggttacgc cggcaaaacc 7800

gatgcgcgcg gcacctacta cggcgagaca gccggttaca ccaccgcgca ggccgaagta 7860

ctgggcaaat atgatagcga aggcaatctc accgccattg gcatctcatt tcgcggcacc 7920

agcggcccgc gcgagtcgtt gatcggcgat accatcggcg atgtgattaa cgatctgctg 7980

gccgggttcg ggccgaaagg ctatgccgaa ggctatacgc tgaaggcctt cggcaatttg 8040

ctgggcgacg tggcgaaatt cgctaaggcc cacgggctga gcggtgagga cgtggtggtc 8100

agcggccaca gcctcggcgg gctggcggtc aatagcatgg cggcgcagag cgacgccaac 8160

tggggcggct tctacgcgca gtccaactat gtcgccttcg cctcgccgac ccagtacgaa 8220

gccggcggta aggtgatcaa catcggctac gagaacgacc cggtgttccg cgcgctcgac 8280

ggcacctcac tgaccctgcc gtcactgggc gtacatgatg cgccgcacgc ctccgccacc 8340

aacaatatcg tcaacttcaa cgaccactac gcgtcggacg cctggaacct gctgccgttt 8400

tccattctca acattccgac ctggctgtcg cacctgccgt tcttttatca ggacggcctg 8460

atgcgggtgc tgaactccga gttttattca ctgaccgaca aggactcgac catcatcgtc 8520

tccaacctgt cgaacgtgac gcgcggcaat acctgggtgg aagacctgaa ccgcaacgcg 8580

gaaacgcaca gcggaccgac gtttatcatc ggcagcgacg gcaatgattt gatcaggggc 8640

ggcaaaggca acgactatct cgagggccgc gacggtgacg atatctttcg cgacgccggc 8700

ggctataacc tgatcgccgg cggaaaaggc cacaatatct tcgataccca gcaggcgttg 8760

aaaaacaccg aggtcgccta cgacggcaac acgctttacc tgcgcgatgc caagggcggc 8820

attacgctgg cggacgacat cagcaccctg cgcagcaaag aaacctcctg gctgattttc 8880

agcaaagagg tggatcacca ggtgacagca accggattga aatcggatgc gggcctcaaa 8940

gcctatgccg ccactaccac cggcggcgac ggcgatgacg tcctgcaggc tcgcggccac 9000

gacgcctggc tgttcggcaa cgccggcaac gacacgctga tcggccacgc cggcggcaat 9060

ctgaccttcg tcggcggcag cggcgatgac atcctgaagg gcgtcggcaa cggcaatacc 9120

ttcctgttca gcggggattt tggccgcgac cagctgtatg gcttcaacgc caccgataaa 9180

ctggtgttta tcggtaccga aggcgccagc gggaatattc gcgactacgc cacgcagcaa 9240

aacgacgatc tggtgctggc cttcgggcac agccaggtca cgctgatcgg cgtctcgctc 9300

gatcacttca ataccgatca ggtggtgttg gcctaaaagc ttgcggccgc actcgagcac 9360

caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct 9420

gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg 9480

ggttttttgc ccaattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc 9540

gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa 9600

gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg 9660

atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc 9720

agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct 9780

gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc 9840

tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag 9900

ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg 9960

tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata 10020

cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataat 10075